Extensive research has been conducted into the role of matrix attachment region (MAR) DNA sequences in the regulation of eukaryotic gene expressions. A MAR sequence (also referred to as a scaffold attachment region (SAR)) is an exemplary element used in the regulation of transcription. In general, a MAR sequence is known to be effective only when inserted into a host genome. It is also known that a MAR sequence, particularly one that is highly rich in AT to an extent of about 70% or greater, increases a transgene expression in an animal cell line that has been stably transformed. It is also known that when a MAR sequence is used, the expression variability of various transformants is low. Such a position-independent expression is believed to be due to the MAR sequence which protects inserted DNA from the intervening effect of neighboring chromatin enhancer or silencer, or inhibits methylation of the inserted DNA, thus insulating foreign DNA inserts from the position effect.
MAR sequences are frequently used to increase expression of foreign genes in animal cells. For example, WO 02/1425 discloses an expression vector containing β-globin MAR sequence at the 5′ terminal of the promoter. U.S. Pat. No. 6,388,066 also discloses a promoter-driven structure containing corn ADH1 MAR DNA sequence which is located adjacent to a combined element consisting of a promoter, a nucleotide sequence operably connected to the promoter, and a transcription termination region. However, a DNA structure having two or more MAR DNA sequences sequentially introduced at the 3′ terminal of a transcription terminator has not been introduced so far.
Even though the technologies as described above are available in the related art, there is still a demand for an expression vector that is capable of expressing foreign genes in animal cells with higher efficiency. While investigating a method of increasing foreign gene expression, the inventors of the present invention have found that foreign gene expression is markedly increased when at least one copy of a MAR DNA sequence is introduced at the 3′ terminal of a transcription terminator of a gene and completed the present invention.